Search PubMed⌕ Search

Biomedical subjects

S Normark

Publications and source records attributed to S Normark.

At least 181 records · Page 10Linked to original sources
PubMed1977

Escherichia coli K-12 mutants hyperproducing chromosomal beta-lactamase by gene repetitions.

Escherichia coli K-12 ampicillin-resistant mutants hyperproducing chromosomal beta-lactamase arose spontaneously from strains carrying ampA1 ampC(+). Such mutants were found even in a recA background. Two Amp(r)-100 strains were analyzed genetically. The Amp(r)-100 resistance level of both strains could be transduced by direct selection for ampicillin resistance. Several classes of ampicillin-resistant transductants were found that differed from one another in the beta-lactamase activity and the ampicillin resistance mediated by an ampA1 ampC(+)-carrying strain. The data suggested that beta-lactamase hyperproduction was due to repetitions of the chromosomal amp genes. The size of the repeated region was calculated from cotransduction estimates, using the formula of Wu (Genetics 54:405-410, 1966), and was found to be about 1 min in one strain and 1.5 min in the other. Second-step Amp(r)-400 mutants were isolated from an Amp(r)-100 strain. The resistance of these mutants was apparently also due to repetitions, each mediating a resistance to about 10 mug/ml. Mutants of wild-type strains that were moderately resistant to ampicillin also gave rise to intermediate-resistance classes, suggesting repetitions of the wild-type amp alleles. F' factors hyperproducing chromosomal beta-lactamase by gene repetitions were constructed. They mediated levels of ampicillin resistance comparable to that of naturally occurring resistance plasmids. The expression of beta-lactamase hyperproduction was not affected by the presence of ampA and ampC alleles in trans and did not act in trans on the other alleles.

Alleles↗
PubMed1976

Septum formation-defective mutant of Escherichia coli.

Mutants of Escherichia coli defective in septum initiation, as well as in septum formation were obtained spontaneously, without mutagenic treatment, by selection of rifampin-tolerant mutants of an antibiotic-permeable strain carrying the envA mutation. The disturbed phenotype was in all mutants aggrevated the low incubation temperatures. One allele, sefA1, was studied in detail. Septum initiation, as well as septum formation, was promoted by high cell densities or by the addition of low concentrations of certain antibiotics, e.g., rifampin and chloramphenicol, to low-density cultures. The observed rifampicin depencence was studied in detail. These experiments indicated that a very modest shift-down situation suppressed the phenotype and enabled constrictions to proceed to cell separation. The rifampicin sensitivity of the partially purified deoxyribonucleic acid polymerase was not affected by the sefA1 allele, which is located close to proA and is thus distinct from envA. Growth parameters during the shift to 25 degrees C were followed in a transductant carrying HE SEFA1 allele. This constriction was characteristically blunt and did not lead to cell separation. At the time of formation of these frozen constrictions, clear zones representing a separation of wall from cytoplasmic membrane appeared. These polar tips did not inhibit expansion of the cell envelope. The phenotype of cells carrying the sefA1 allele suggests a disturbed relationship among protoplasm expansion, envelope growth, and septum formation. It is thought that the blunt constrictions observed are caused by an inability of the two septal peptidoglycan layers to fuse during an early stage of septation.

Cell Division↗
PubMed1976

Evidence for a role of N-acetylmuramyl-L-alanine amidase in septum separation in Escherichia coli.

Septum formation and septum separation have been studied in a chain-forming mutant of Escherichia coli K-12 bearing the envA mutation and its parental strain. In comparison to the wild type, the mutant showed a sixfold reduction in the specific activity of the enzyme, N-acetylmuramyl-L-alanine amidase (EC 3.5.1.28), part of which was associated to the outer membrane. Genetic as well as physiological suppression of chain formation resulted in an increase in amidase activity. The addition of N-acetylmuramyl-L-alanyl-D-glutamyl-meso-diaminopimelic acid to growing wild-type cells and to cells bearing the envA mutation caused an inhibition of cell separation and an increased frequency of visible septa. The kinetics of septum formation and separation was followed in chains by the use of ampicillin and nalidixic acid. The latter drug inhibited initiation of new septa but allowed preformed ones to go to cell separation at a rate corresponding to that of steady-state growing cells. Ampicillin treatment, on the other hand, resulted in a more rapid decrease in the frequency of septa. The disparate effects of ampicillin and nalidixic acid were not explained by a difference in amidase activity but could be due to an inhibitory effect of ampicillin on a septal peptidoglycan fusing activity.

Amidohydrolases↗
PubMed1976

Scanning electron microscopy of Neisseria gonorrhoeae. Age-induced changes in macro- and microstructure of virulent and avirulent colonies.

Virulent and avirulent gonococcal colonies were studied in the scanning electron microscope. Cells in virulent, in contrast to avirulent, colonies were found to be connected by numerous strands. After longer incubation periods, type 2 colonies segregated out regions of cells virtually lacking these strands. Cells within these areas probably represent avirulent segregants. The microstructure of individual colonies revealed no topographical differences. In certain colonies large smooth spherical cells were seen, probably representing wall-deficient gonococcal variants.

Microscopy, Electron, Scanning↗
PubMed1976

Cell envelope of Neisseria gonorrhoeae. A comparative study with Escherichia coli.

The cell envelope of Neisseria gonorrhoeae was studied and compared to that of Escherichia coli. Outer membranes (OM) from both species were isolated by an identical method, and subjected to biochemical analysis. Differences in OM structure were sought that might explain the dissimilarity in OM permeability of these two species. The most pronounced difference appeared to reside in the OM proteins as judged by gel electrophoresis. Moreover, gonococcal OM proteins appeared to be more hydrophilic than those of E. coli.

Bacterial Proteins↗
PubMed1975

Cell envelope of Neisseria gonorrhoeae: outer membrane and peptidoglycan composition of penicillin-sensitive and-resistant strains.

The cell envelope of Neisseria gonorrhoeae, colony type 4, was studied. Outer membrane was isolated by lysozyme and ethylenediaminetetraacetic acid treatment of plasmolyzed cells according to Wolf-Watz et al. (1973). The degree of purity of the membrane preparations was checked by electron microscopy. The membrane fraction obtained had a density of 1.25 g/cm(3), was rich in phospholipase A and lysophospholipase, and contained only 10% of the total membrane activity of succinate dehydrogenase and d-lactate dehydrogenase. The outer membrane protein profile after sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed at least six major proteins. The predominating protein showed a molecular weight of 35,000. The lipopolysaccharide component was characterized by gas chromatography. The carbohydrates found were galactose, glucose, and glucosamine. d-Glycero-l-manno-heptose was present in very low amounts. Lipid A contained lauric acid, stearic acid, and beta-hydroxy-myristic acid. About 20% of the fatty acids in the outer membrane was derived from lipid A. The phospholipids were characterized as phosphatidylethanolamine, phosphatidylglycerol, and diphosphatidylglycerol. There was no evidence for a lipoprotein anchored to the peptidoglycan. The peptidoglycan of N. gonorrhoeae was of the chemotype I. The cell envelope of N. gonorrhoeae was found to be highly permeable to gentian violet. Cell envelopes of one penicillin-resistant and two penicillin-sensitive strains were compared. Only moderate differences in fatty acid composition were found.

Carbohydrates↗
PubMed1975

Genetic and physiological analysis of an envB spherelike mutant of Escherichia coli K-12 and characterization of its transductants.

The envB1 mutation mediating a distorted cell morphology of Escherichia coliK-12 was cotransducible with strA, aroE, aspB, and argG. The mapping data is consistent with a gene location for envB around 62.5 min. In partial diploids envB1 was recessive to its wild-type allele. The original envB mutant contained a second mutation in a locus denoted sloB close to strA. The following gene order is suggested: sloB-strA-aroE-envB-aspB-argG. The sloB1 mutation caused a marked reduction in the growth rate of both envB and envB+ strains. Moreover, this mutation in the presence of envB1 appears to increase the ratio between deoxyribonucleic acid and protein in cells growing in rich medium. The phenotypic properties of envB1, sloB+ and envB+ transductants were characterized. Cells with envB1, sloB+ genotype were hypersensitive to several penicillins including the beta-lactam compound, amidino penicillin. Penicillin hypersensitivity could not be explained by increased outer membrane penetrability. The original envB mutant (envB1,SLOB1), as well as envB1, sloB1 or envB+, SLOB1 transductants were resistant to amidino penicillin. Resistance was explained by the slow growth rate mediated by the sloB1 mutation. The similarity between envB cells and wild-type cells treated with sublethal concentrations of amidino penicillin was emphasized.

Bacterial Proteins↗
PubMed1973

Rapid method for isolation of large quantities of outer membrane from Escherichia coli K-12 and its application to the study of envelope mutants.

A rapid method for the isolation of large quantities of bacterial outer membrane is described. This cell envelope component was removed from plasmolyzed cells of Escherichia coli K-12 by lysozyme-ethylenediaminetetra-acetic acid treatment, aggregated by lowering the pH to 5.0, and recovered by centrifugation. Aggregates of membrane fragments were clearly identified in an electron microscope. A criterion of homogeneity of the preparation was obtained by isopycnic sucrose gradient centrifugation. A single band appeared at a density of 1.24 g/cc. The cytoplasmic membrane marker, succinate dehydrogenase activity, was 40 times lower in the outer membrane preparation than in complete cell envelope preparations. A rich activity was, however, found for the outer membrane marker, phospholipase A. The compositions of outer membranes from a transductant pair were compared. One transductant was a chain-forming, antibiotic-supersensitive envA strain, whereas the other contained the envA(+) allele. The envA strain showed a slightly modified protein pattern and a lower relative content of phosphatidylglycerol.

Bacterial Proteins↗
PubMed1973

Outer penetration barrier of Escherichia coli K-12: kinetics of the uptake of gentian violet by wild type and envelope mutants.

Wild-type strains of Escherichia coli K-12 adsorb gentian violet to the cell surface, but the dye is not transported into the cytoplasm. However, when some mutants that have an altered outer membrane are exposed to gentian violet, the dye is also found in the ribosomal fraction. The transport into the cytoplasm is inhibited at 0 C and requires that the concentration of gentian violet exceeds a threshold value. The initial rate of uptake as well as the amount of gentian violet found in the cytoplasm increases with the concentration of the dye in the medium. The rate of transport of the dye into the cytoplasm is much lower for stationary mutant cells than for exponentially growing cells. The rate of uptake into the cytoplasm increases with increasing deficiency of carbohydrate in the lipopolysaccharide (carbohydrate content lpsB > lpsA > galU). However, other components are also responsible for the barrier since an envA mutant which is not altered in the lipopolysaccharide carbohydrates show an extremely rapid uptake of the dye. The rate of uptake for the envA mutant was the highest found and the same as that of spheroplasts. Growth in the presence of agents affecting the murein sacculus, e.g., lysozyme and sublethal concentrations of penicillin, increased the rate of uptake of gentian violet. Brief treatments with tris(hydroxymethyl)aminomethane-ethylenediaminetetraacetic acid drastically impaired the barrier function. Inhibition of protein synthesis by chloramphenicol also opened the barrier to gentian violet. In conclusion, the outer part of the bacterial envelope is a penetration barrier for gentian violet and probably also for other substances. The lipopolysaccharide, the murein and also other components are important for the function of this barrier. Resistance to gentian violet was found to be inversely correlated to the rate of penetration of the dye into the cytoplasm.

Bacterial Proteins↗
PubMed1972

In vitro studies of 5-fluorocytosine resistance in Candida albicans and Torulopsis glabrata.

Spontaneous mutants of Candida albicans resistant to 5-fluorocytosine (5-FC) were isolated from a strain susceptible to 5-FC. These mutants were compared with 5-FC-resistant strains of C. albicans and Torulopsis glabrata isolated from patients treated with the drug and from untreated patients. Resistance to 5-FC was in all cases followed by a decreased susceptibility to 5-fluorouracil. Most strains were also more resistant to 5-fluorouridine and had a lowered incorporation of uridine. In one spontaneous mutant totally resistant to all three 5-fluoropyrimidines, the uridine monophosphate pyrophosphorylase activity was greatly decreased. The 5-FC-resistant strains were of two main phenotypic classes. Class one was unaffected by 5-FC at the highest concentration tested. The growth rate of strains belonging to the other class was markedly decreased by low concentrations of 5-FC, but these strains were still able to form colonies after 7 days of incubation on plates containing very high concentrations of the drug (2,000 mug/ml). The frequency of spontaneous mutation of a susceptible strain to 5-FC resistance was fairly high. The number of mutants growing on media containing 5-FC was unchanged by concentrations ranging from 25 to 500 mug/ml.

Antifungal Agents↗
PubMed1971

Nature of the penetration barrier in Escherichia coli K-12: effect of macromolecular inhibition of penetrability in strains containing the envA gene.

The envA mutation in Escherichia coli K-12, which maps at 1.5 min, was previously shown to mediate sensitivity to gentian violet as well as to several antibiotics. Moreover, strains containing the envA gene were recently found to be lysed by lysozyme in the absence of ethylenediaminetetraacetate. It is here reported that the envA mutation mediates an increased uptake of gentian violet. The uptake of the dye was markedly affected by growth with different antibiotics interfering with macromolecular synthesis. Amino acid starvation of a strain containing envA with a stringent control of ribonucleic acid (RNA) synthesis resulted in a decreased uptake of gentian violet. However, no decrease in dye uptake was found during starvation in an envA transductant with a relaxed control of RNA synthesis. Inhibition of deoxyribonucleic acid (DNA) synthesis by nalidixic acid decreased the uptake of gentian violet of envA cells and, in addition, rendered the cells insensitive to the lytic action of lysozyme. Chloramphenicol treatment increased penetrability in wild-type and starved envA cells. In most instances, this effect of chloramphenicol was prevented by selectively interfering with DNA or RNA synthesis. A coordinate regulation of nucleic acid synthesis and penetrability is suggested.

Amino Acids↗
PubMed1971

Phenethyl alcohol as a suppressor of the envA phenotype associated with the envA gene in Escherichia coli K-12.

In Escherichia coli K-12 the envA gene was previously shown to mediate chain formation and a decreased tolerance to several antibacterial agents. Phenethyl alcohol at low concentrations has now been found to increase the tolerance to actinomycin D, ampicillin, rifampin, and gentian violet in strains containing envA. The increased tolerance to gentian violet was correlated to a decreased uptake of the dye. A phenotype suppression of chain formation and colony morphology in envA mutants was also obtained. Except for an increase in palmitic acid, chemical analysis revealed no differences between an envA and its wild-type strain in the lipopolysaccharide part of the envelope. However, a decrease in the amount of phosphatidylglycerol and a C18: 1 fatty acid was observed in the extractable lipids of a strain containing envA. Growth in the presence of phenethyl alcohol reversed the changes in fatty acid and the phospholipid composition. Phenethyl alcohol was found to cause an immediate but transient inhibition of ribonucleic acid synthesis. It is suggested that this inhibition affects the penetrability barrier of the outer cell envelope layers in strains containing envA.

Alcohols↗